The cardiac sarcolemmal Na-Ca exchanger (NCX) is the primary mechanism for the extrusion of Ca from myocytes. As such, the exchanger is an important regulator of intracellular Ca and cardiac contractility. The long-term objective of this application is to further understand the significance of the Na-Ca exchanger in excitation-contraction (EC) coupling. Genetic alterations in the level of the Na-Ca exchanger have had unexpected effects on EC coupling. The two proposed projects are as follows: 1. Adaptations of cardiac EC coupling to genetically altered levels of Na-Ca exchanger. Mice with large changes in the level of the Na-Ca exchanger have been created in the laboratory of the PI. The most interesting of these mice are transgenic homozygous NCX overexpressors (>3-fold increase in activity) and cardiac-specific NCX knockout (KO) mice. The NCX KO mice unexpectedly live to adulthood and demonstrate a surprising plasticity of EC coupling pathways. The myocardium adapts to the absence of the primary Ca efflux mechanism by decreasing Ca influx into myocytes by 80%. Experiments are proposed to explore the contributions of various mechanisms to this adaptation. 2. A biochemical correlate of altered gain of EC coupling. Overexpression of the sarcolemmal Na-Ca exchanger induces a change in the membrane environment of the dihydropyridine receptor (DHPR or L-type Ca channel);the DHPR is located in lipid rafts in wild type myocytes, but the DHPR no longer fractionates in lipid rafts in NCX-overexpressing myocytes. Correlating with this re-distribution is a decrease in the gain of EC coupling. We hypothesize that overexpression of NCX changes the physical relationship between DHPRs and ryanodine receptors which leads to diminished coupling between these proteins and a decrease in gain. Understanding how NCX interacts with the EC coupling machinery is essential to understanding the altered Ca fluxes in heart failure and other pathologies. NCX has unexpected regulatory effects on EC coupling with pharmacological implications.